Abstract
Cisplatin (CisPt) is frequently used in the therapy of urothelial carcinoma (UC). Its therapeutic efficacy is limited by inherent or acquired drug resistance. Here, we comparatively investigated the CisPt-induced response of two different parental urothelial carcinoma cell lines (RT-112, J-82) with that of respective drug resistant variants (RT-112R, J-82R) obtained upon month-long CisPt selection. Parental RT-112 cells were ~2.5 fold more resistant to CisPt than J-82 cells and showed a different expression pattern of CisPt-related resistance factors. CisPt resistant RT-112R and J-82R variants revealed a 2–3-fold increased CisPt resistance as compared to their corresponding parental counterparts. Acquired CisPt resistance was accompanied by morphological alterations resembling epithelial mesenchymal transition (EMT). RT-112R cells revealed lower apoptotic frequency and more pronounced G2/M arrest following CisPt exposure than RT-112 cells, whereas no differences in death induction were observed between J-82 and J-82R cells. CisPt resistant J-82R cells however were characterized by a reduced formation of CisPt-induced DNA damage and related DNA damage response (DDR) as compared to J-82 cells. Such difference was not observed between RT-112R and RT-112 cells. J-82R cells showed an enhanced sensitivity to pharmacological inhibition of checkpoint kinase 1 (Chk1) and, moreover, could be re-sensitized to CisPt upon Chk1 inhibition. Based on the data we suggest that mechanisms of acquired CisPt resistance of individual UC cells are substantially different, with apoptosis- and DDR-related mechanisms being of particular relevance. Moreover, the findings indicate that targeting of Chk1 might be useful to overcome acquired CisPt resistance of certain subtypes of UC.
Highlights
Bladder cancer is a frequent type of cancer worldwide
Based on the data we suggest that mechanisms of acquired CisPt resistance of individual urothelial carcinoma (UC) cells are substantially different, with apoptosis- and DNA damage response (DDR)-related mechanisms being of particular relevance
Based on a recent report of Galluzzi et al [17], who has classified putative CisPt resistance factors of tumor cells, we assembled a 96 well-based quantitative real-time PCR array to comparatively analyze the mRNA expression of these factors in reverse transcriptase (RT)-112 and J-82 cells
Summary
Bladder cancer is a frequent type of cancer worldwide. In most countries, the majority of bladder cancers are urothelial carcinoma (UC) [1]. Upon replacement of its chloride ligands by water, DNA adducts are formed by SN2 mechanism (nucleophilic substitution) [5]. The vast majority (60–80%) of DNA adducts generated by CisPt are DNA intrastrand crosslinks (GpG and ApG). Platinuminduced DNA crosslinks cause a substantial distortion of the DNA double helix, resulting in transcription and replication blockage [8, 9]. In consequence of stalled replication forks, DNA double-strand breaks (DSBs) can arise as secondary lesions [10]. DSBs are potent triggers of cell death [11] and can be repaired by DNA doublestrand break repair (homologous recombination (HR) or non-homologous end joining (NHEJ)). The removal of CisPt-induced DNA crosslinks involves nucleotide excision repair (NER), including transcription-coupled www.impactjournals.com/oncotarget
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